Medulloblastoma is one type of primitive neuroectodermal tumors which account for 40% of all brain tumors with glioma, and is a malignant brain tumor that frequently appears in children under 10 years of age, and particularly between the ages of 5 to 10 years. This tumor is a typical undifferentiated tumor that mimics morphology and arrangement patterns of undifferentiated cells that compose the embryonal medullary epithelium and matrical layer. It is considered to be an immature tumor that has the potential to differentiate into both nerve cells and glia cells (Tamura, K. et al., Cancer Res., 49, 5380-5384 (1989)). Since these types of malignant tumors exhibit sensitivity to radiation and chemotherapeutic agents, it is treated by radiotherapy, chemotherapy as well as surgery.
However, although these treatment methods alleviate symptoms temporarily, there are many cases of relapse and death within several years, with an average survival period being 15 months. The cause of this is believed to be that a recurrent cancer has resistance to chemotherapy and radiation.
On the other hand, accurate evaluation of brain tumors relies heavily on histological techniques, and requires an extremely high level of specialized knowledge as well as auxiliary diagnostic technology.
Thus, there is a pressing need for the development of diagnostic tools and therapeutic drugs that enable early diagnosis and fundamental treatment.
Several attempts have been made at treatment through the use of monoclonal antibodies that recognize medulloblastoma in the past as well (refer to Kemahead, J. T. et al., Int. J. Cancer, 31, 187-195 (1983), Allan, P. M. et al., Int. J. Cancer, 31, 591-598 (1983), Jones, D. et al., Br. J. Hematol., 57, 621-631 (1984), Gross, N. et al., Cancer Res., 46, 2998-2994 (1986), Wikstrand. C. D. et al., Cancer Res., 46, 5933-5940 (1986), Gibson, F. M. et al., Int. J. Cancer, 39, 554-559 (1987), Feickert, H. J. et al., Cancer Res., 49, 4338-4343 (1989), Jennings, M. T. et al., J. Neurol. Sci., 89, 63-78 (1989) and Takahashi, H. et al., Neurosurg., 27, 97-102 (1990)).
However, since nearly all of these antibodies also recognize normal tissue and other tumors, they have the disadvantage of being inappropriate for diagnosis and treatment of medulloblastoma.
Recently, brain tumor immunotherapy has been reported that uses a monoclonal antibody of human origin that reacts to the human glioma and exhibits ADCC activity (see, Japanese Unexamined Patent Publication No. 58-201994, Japanese Unexamined Patent Publication No. 59-137497 and Japanese Unexamined Patent Publication No. 4-356792).
Mouse monoclonal antibody exhibits a high degree of immunogenicity (also referred to as antigenicity) in humans. For this reason, their therapeutic value in humans is limited. Moreover, not only do mouse antibodies inhibit anticipated effects, but they cannot be administered frequently without provoking an immune reaction that brings about the risk of an allergic response that presents a problem for patients.
In order to solve these problems, a process for producing humanized antibody has been developed. Mouse antibody can be humanized by two methods. The simpler method involves the production of chimeric antibody wherein the variable region is derived from an original mouse monoclonal antibody, while the constant region is derived from a suitable human antibody. The resulting chimeric antibody contains a complete variable region of the original mouse antibody, and can be expected to bind antigen with the same specificity as the original mouse antibody.
Moreover, since the ratio of the protein sequence derived from sources other than humans is essentially reduced, it is predicted to have a low level of immunogenicity in comparison with the original mouse antibody. Although chimeric antibody effectively binds with antigen and has a low level of immunogenicity, there is still possibility of an immune reaction to the mouse variable region (LoBuglio, A. F. at al., Proc. Natl. Acad. Sci. USA, 86, 4220-4224, 1989).
Although more complex, the second method for humanizing mouse antibody considerably lowers the potential immunogenicity of the mouse antibody even more. In this method, complementarity determining regions (CDRs) from variable regions of a mouse antibody are transplanted into human antibody variable regions to produce a reshaped human antibody variable regions.
Next, these reshaped human antibody variable regions are linked to human antibody constant regions. Ultimately, the portion derived from the non-human protein sequence of a reshaped human antibody is only the CDRs and an extremely small portion of the framework (FR). CDRS comprise hyper-variable protein sequences. These sequences do not exhibit type-specific sequences. For these reasons, a reshaped human antibody containing mouse CDRs ought not to have immunogenicity stronger than naturally-occuring human antibody containing human CDRs.
The following references should be referred to with respect to reshaped human antibodies: Riechmann, L. et al., Nature, 332, 323-327, 1988; Verhoeyen M. et al., Science, 239, 1534-1536, 1988; Kettleborough, C. A. et al., Protein Engng., 4, 773-783, 1991; Maeda, H. et al., Human Antibodies and Hybridoma, 2, 124-134, 1991; Gorman S. D. et al., Proc. Natl. Acad. Sci. USA, 88, 4181-4185, 1991; Tempest P. R. et al., Bio/Technology, 9, 266-271, 1991; Co, M. S. et al., Proc. Natl. Acad. Sci. USA, 88, 2869-2873, 1991; Carter, P. et al., Proc. Natl. Acad. Sci. USA, 89, 4285-4289, 1992; Co, M. S. et al., J. Immunol., 148, 1149-1154, 1992; and, Sato, K. et al., Cancer Res., 53, 851-856, 1993.
As was previously stated, although it is predicted that a reshaped human antibody is useful for the purpose of therapeutical treatment, a reshaped human antibody to human medulloblastoma cells is not known. Moreover, there are no methods for producing a reshaped human antibody that can be universally applied to any specific antibody. Thus, various contrivances are necessary to produce a reshaped human antibody to a specific antigen that has sufficient activity (for example, Sato, K. et al., Cancer Res., 53, 1-6 (1993)).
The inventors of the present invention isolated and established a medulloblastoma cell line (ONS-76) from the cerebellum of medulloblastoma patients (Tamura, X. et al., Cancer Res., 49, 5380-5384 (1989)). By then immunizing mice with said medulloblastoma cell line ONS-76, a mouse monoclonal antibody (ONS-M21) was found that specifically recognizes human medulloblastoma but does not cross-react with normal brain tissue or peripheral blood cells (Moriuchi. S. et al., Br. J. Cancer, 68, 831-837 (1993)). Since antigen recognized by this antibody is strongly expressed in brain tumors such as medulloblastoma and some gliomas, it is anticipated to be used as a diagnostic tool as well as directly destroy cancer cells by inducing ADCC and CDC or conjugating with toxins and radioisotopes.